V1534 000 r m h

Mice were treated as described previously [32] (link). In short, 52 week (wk) old C57BL/6J mice (Charles River Laboratories, Wilmington, MA) were exposed to ambient air (normoxia, n = 8) or chronic hypoxia (n = 7) for 21 days. To control for the effects of reduced food intake during hypoxia, another group of mice was exposed to ambient air while receiving the amount of food consumed by the hypoxic mice (pair-fed group, n = 8). All mice were housed in experimental chambers at 21°C with a 12-h dark/light cycle. Mice received standard chow (V1534–000 ssniff R/M-H, ssniff Spezialdiäten, Soest, Germany) and water ad libitum. Using the proOX P110 (BioSpherix, Lacona, NY) system, O2 was replaced by N2 in a stepwise manner to create normobaric oxygen levels of 12% (day 1), 10% (day 2), and finally 8% (60.8 mmHg) on day 3. The latter oxygen concentration was maintained until day 21. Three to four mice were housed per cage. Daily food intake was determined per cage, and mice were weighed daily. On day 21, mice were sacrificed. The lungs were snap frozen in liquid nitrogen and stored immediately in −80°C. The protocol was approved by the Committee for Animal Care and Use of Maastricht University (project 2009-151).

All mice were maintained under conventional conditions and a 12:12 h light:dark cycle (lights on from 6 a.m. to 6 p.m.) at a temperature of 22 ± 2 °C. Animals had ad libitum access to food and water. The mice were fed with a rodent standard diet (V1534-000 ssniff R/M-H; Ssniff Spezialdiäten GmbH, Soest, Germany).

All experimental treatments of animals were authorized by the German Animal Welfare Authorities (approval no. G0174/19). All methods were performed in accordance with the relevant guidelines and regulations of German Animal Welfare laws and also in accordance with Animal Research: Reporting of In Vivo Experiments (ARRIVE) guidelines. All mice were maintained under conventional conditions and a 12:12 h light: dark cycle at a temperature of 22 ± 2 °C. Animals had ad libitum access to food (rodent standard diet (V1534-000 ssniff R/M-H; ssniff Spezialdiäten GmbH, Soest, Germany)) and water.